Project Summary Vascular homeostasis and function is reliant upon intercellular communication with and within the vascular wall. Perturbations in homeostatic signaling can lead to development of cardiovascular disease, such as atherosclerosis, hypertension, and heart disease. Recently, small nanovesicles termed `exosomes', have been described as a novel means of communication between cells. Specifically, exosomes are 30-100nm vesicles that are packaged within the cell and released into extracellular space upon fusion with plasma membrane. Exosomes contain unique cargo of proteins, such as transmembrane receptors and cytosolic signal transduction proteins, in addition to small RNAs, such as microRNAs and non-coding RNAs, all of which are capable of initiating signal transduction and altering gene expression in target cell. Despite the well- characterized role in immunology and cancer biology, little is known on the role of exosomes in vascular biology. The vascular artery consists of a confluent layer of endothelial cells (ECs) surrounded by layers of contractile smooth muscle (VSMC). Inflammation or insult to either layer disrupts homeostasis and can lead to disease. The proposed research will focus on the signaling mechanisms between the endothelial and smooth muscle cell layers through exosome-mediated transfer. Our preliminary data strongly suggests that endothelial exosomes can initiate changes in protein expression in VSMCs leading to alteration of function. The long-term goal of this research is to elucidate the mechanisms through which exosomes signal between cells in the vascular wall and under what conditions they may be protective or pathogenic. Of particular interest is the method through which exosomes signal once they reach recipient cell. We will test the hypothesis that endothelial exosomes mediate vascular inflammation and remodeling through AP-1/NF-kB activation and concurrent mitochondrial fragmentation through the following specific aims: 1) Identify the signaling mechanism and downstream consequences of EC exosomes in VSMCs in vitro, 2) Test the signaling mechanism and downstream consequences of EC exosomes in ex vivo organ culture, and 3) Elucidate the effects of EC exosomes on neointimal formation in an in vivo model of rat carotid balloon injury. With the excellent tutelage of my mentors and an environment to excel, this proposal will elucidate key findings in the understanding of vascular pathophysiology. The analytical and technical training provided through the fellowship will promote my development as an independent scientist and serve as a critical step moving forward in my career.